The construction and application of adenovirus vector coexpressing the heterodimer of human IL 12

Zhang Weiping,Cao Xuetao (Department of Immunology, Second Military Medical University, Shanghai 200433)  

Objective To construct the recombinant adenovirus vector co expressing the heterodimer of human interkeukin 12.Methods The full length cDNA encoding human IL 12 subunits p40 or p35 was cloned by RT PCR separately. The cDNA was ligated with encephalomyocarditis internal ribosome entry site (IRES), placed under the control of CMV promoter, and inserted into E1 substituted adenovirus vector pAx1cw to produce the bicistronic coexpression vector. Subsequently, the hIL 12 recombinant adenovirus vector was cotransfected into 293 cells together with EcoT22I digested Ad5 DNA TPC,and the replication deficient hIL 12 recombinant adenovirus was generated efficiently by homologous recombination.Results The human IL 12 recombinant adenoviruses were obtained with the titers of 2.1×10 9pfu/ml.48 hours after the infection of the 293 cells, HepG2 cells and huaman primary skin fibroblasts with hIL 12 recombinant adenoviruses, the hIL 12 expressions were detected by ELISA (30～50ng/10 6cells/24h). The expressed hIL 12 could stimulate the in vitro proliferation and IFN γ production of human PMNC.Conclusion Prepared hIL 12 recombinant adenovirus vector can express biologically active hIL 12 and can be potentially used in cancer gene therapy.

【Key Words】： Interleukin 12 Adenovirus Gene expression
【Fund】： 国家自然科学基金 优秀中青年人才专项基金
【CateGory Index】： R373,

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